PROJECT SUMMARY The proposed studies in lampreys and hagfish build on the findings that both jawless (agnathans) and jawed (gnathostomes) vertebrates have prototypic T-like and B-like lymphocyte lineages, although jawless vertebrates (lampreys and hagfish) generate variable lymphocyte receptors (VLR) for antigen recognition by recombinatorial conversion of incomplete germline VLR genes (VLRA, VLRB and VLRC) into fully assembled VLR genes using diverse leucine-rich-repeat (LRR) donor sequences. Analysis of the development, distribution and function of the VLRA and VLRC lymphocyte lineages suggest that the VLRA and VLRC lineages represent the agnathan T cell analogues of gnathostome ?/? and ?/? T cells with allorecognition responsibility. The multistep assembly of VLRC and VLRA genes coincides with expression of cytidine deaminase1 (CDA1) in a thymus-equivalent region of the gills, whereas VLRB and CDA2 expression occurs primarily in hematopoietic tissues. Surprisingly, three additional CDA1-like genes, the products of which have deaminase and mutagenic activity, have now been identified. The proposed studies will define the newly discovered multigene CDA1 family members in agnathans, determine their cellular expression patterns and elucidate the structure and functions of their protein products. Circumstantial evidence implies that the VLRA and VLRC receptors rely on associated transmembrane molecules for their cell surface expression and signaling competence. The present studies will determine the composite receptor structure and signaling competence for VLRA and VLRC. Pilot studies indicate that the VLRA+ and VLRC+ lymphocytes preferentially respond to allogeneic white blood cells. The antigens recognized by VLRA and VLRC receptors will be defined with the related goal of identifying lamprey histocompatibility antigens and determining their roles in antigen presentation. Candidate histocompatibility genes will be examined for genetic polymorphism, expression patterns and immunostimulatory potential to test the hypothesis that VLRA and VLRC recognize histocompatibility-associated antigens. Identification of the agnathan histocompatibility genes will enlighten us about the mechanisms of self versus non-self discrimination at a pivotal juncture of vertebrate evolution.